Infertility affects approximately 1.2 million married couples in the United States and many more worldwide; of those around 50% of the infertility problems are fully or partially attributed to male-related factors which include: low sperm count, abnormal spermatogenesis, and reduced androgen production. The incidence of male infertility cases has increased and the reasons for the increase in testicular abnormalities are not well understood but may be a direct result of the disruption of cell differentiation. Spermatogonial stem cells (SSCs) are essential for spermatogenesis because the population of SSCs differentiates to produce spermatogonia. Remarkably, despite the importance of SSCs for continual production of sperm, scientists have yet to definitively identify factors that directly regulate SSC differentiation and proliferation. Thus, the identification of factors that regulate SSCs is expected to provide a better understanding of testes development and contribute to potential treatments for male infertility caused by improper cell differentiation. The long-term goal of this research is to elucidate cellular and molecular mechanisms regulating SSC differentiation and proliferation. The objective of this proposal, the first step toward attaining our long-range goal, is to determine the role vitamin A plays in regulating SSC differentiation and self-renewal. The central hypothesis of this proposal is that vitamin A is critical for the differentiation and self-renewal of SSCs in the testes. We will test the effect of vitamin A on SSCs directly by treating SSCs in culture with retinoic acid, the active metabolite of vitamin A. The rationale that underlies the proposed research is that identification of factors that regulate SSCs will lead to mechanistic studies that will provide a better understanding of germ cell abnormalities leading to infertility in males. To test the hypothesis and accomplish the overall objective of this application 2 specific aims are proposed. Specific Aim 1: Determine if vitamin A regulates the conversion of gonocytes to SSCs. Specific Aim 2: Determine if vitamin A regulates the proliferation and self-renewal of SSCs. The proposed research is innovative, because it increases the knowledge of cellular and molecular mechanisms that regulate SSCs. The current proposal is novel because this is the first time that the effect of a factor, such as vitamin A, on SSC differentiation has been evaluated. [unreadable] [unreadable] [unreadable]